Method for producing combination immunometabolic preparation with anti-infectious activity

FIELD: medicine.

SUBSTANCE: group of inventions refers to veterinary science and aims at normalising metabolic processes, stimulating immune system and blocking mechanisms of infectious process with a risk of endogenous infection activation. A method for producing a combination immunometabolic preparation with anti-infectious activity involves dissolving succinic acid and levamisole in demineralised water with formalin added. According to the other aspect of the invention, the immunometabolic preparation additionally contains polyethylene glycol. The ingredients are used in the declared ratio.

EFFECT: using this group of inventions provides producing an injection form of the immunometabolic preparation with anti-infectious activity.

2 cl, 3 tbl

 

The invention relates to veterinary medicine and relates to a method of obtaining a complex of the drug on the basis of succinic acid, levamisole, formalin and polyethylene glycol, which is intended to normalize the metabolism, stimulate the immune system and blocking mechanisms in the development of infectious process, at the risk of activation of endogenous infection, including for the treatment of infectious diseases of viral and bacterial etiology.

A method of obtaining drug "amber biostimulator" to increase the resistance of the organism of animals (RF patent No. 2303979, 2007). The combination of succinic acid and antiseptic stimulant Dorogov second fraction (SDA-f No. 2) provides the ability to get the drug with pronounced immunometabolic activity. However, succinic acid will neutralize the antiseptic activity of the SDA-f No. 2, which ensured a high alkaline reaction.

As the closest analogue of the selected method of obtaining immunotropic antiseptic drug for the treatment and prevention of infectious animal diseases (patent RF №2361579, 2009). This drug contains succinic acid, SDA-f # 2 and formalin (formal amber biostimulant).

In addition immunometabolic drug has a pronounced antiseptic activity. But not what atcom this drug is used as immunostimulant DCA-f No. 2. ASD f-No. 2 obtained by high temperature sublimation of meat and bone meal. The resulting product has a highly variable chemical composition, which complicates its use as a medicinal substance. Also used for the manufacture of the SDA-f No. 2 raw - meat-bone meal can contain high amounts of heavy metals, including radionuclides that accumulate during the life of animals in the bone tissue.

The task of the invention to provide injectable form immunometabolic drug with anti-infective activity.

Serves as immunostimulant to include in the proposed standard drug substance of levamisole.

Levamisole has long been known and are widely used in medicine to stimulate the cellular immune system. In veterinary medicine it is used as an anthelminthic drug. However, levamisole has a strong side, mainly convulsive effect on the body.

What's new is that the integrated product includes in its composition as immunostimulant levamisole; stabilizer and prolongator - polyethylene glycol (PEG).

The combination of metabolica (succinic acid), immunomodulator of levamisole and formalin is quite acceptable due to the different mechanism of action.

Formalin is about the fully exceptionally high antiseptic activity. So, in a concentration of 1:6000 it stops the growth of the typhoid bacilli, and absolutely low concentration of 1:30000 stops the rot broth.

According to Lutcavage NR. (1997), intramuscular 0.2% formalin allows to reduce the virulence of the pathogen virus transmissible gastroenteritis and simultaneously to stimulate specific immune protection. Formaldehyde has a protective effect on the living cells of organs and tissues, inhibits the damaging processes of lipid peroxidation, is present in a normally functioning tissue, can prolong the time of death, heart and brain.

The above effects have formed the basis for inclusion of formaldehyde in the composition of our previously developed immunometabolic drug - amber biostimulant (RF Patent No. 2303979).

The inclusion of the drug PEG provides stability and prolongation of action. It is very important that the glycols with a molecular weight of up to 600 can be mixed with water in any relationship and have a pronounced low toxicity. In pharmacology, the glycols are considered practically non-toxic substances. Currently, the glycols are widely used in pharmaceutical, cosmetic, food industry.

In preliminary experiments nabilah mice was established, that intraperitoneal injection in a volume of 0.25 ml, comprising a standard 10% solution of levamisole, which added a 1% succinic acid, caused a pronounced low toxicity on the body. An even more pronounced reduction of side effects was observed when adding to 1.5 and 2% succinic acid 10% solution of levamisole. In turn, the composition comprising from 1% to 2% succinic acid and 2-3% of levamisole, did not cause any side effects on the body with three times with an interval of 24 hours intraperitoneal administration. Moreover, experimental animals (white mice) became more cheerful, and they had improved appetite.

This object is achieved by inclusion in the composition of the complex preparation of succinic acid, levamisole, formaldehyde and glycol in the following ratio, wt.%:

Succinic acid is 1.0 to 1.5

Levamisole - 2-3

Formalin - 0,3-0,4

The glycol - 7-8

Example 1 of the method of sample preparation of the drug.

For preparation of the comprehensive drug is used in 950 ml of demineralized water, which when heated successively dissolved 10.0 g of succinic acid and 20 g of levamisole (1st version); 15 g of succinic acid and 30 g of levamisole (2nd option). In the cooled to 40-50°C. the composition of each sample was added 10.0 ml of formalin (39% concentration). Add in the solutions is poliatilenglikola to 7-8% increased their density. The total volume was brought to 1000 ml by adding demineralized water. The resulting solution had a pH of 4.5 to 4.7.

In a similar manner there were prepared samples of similar compositions, but without the addition of polyethylene glycol.

Sterilization by autoclaving did not change the transparency of the solution, which testified to the full compatibility of the components.

Test safety. Experiments conducted on white mice. Daily, for 3 days intraperitoneal injection in a volume of 0.25 ml of the above samples of the test drug was not accompanied by the suppression of the General condition experimental animals. On the contrary, the introduction of levamisole in similar concentrations was accompanied by a severe depression lasting from 20 to 30 minutes to 2-3 hours.

The effectiveness of the proposed drug tested in tests of anti-infective and immunometabolic actions.

The test of protective properties

Object for the experiments served as white mice with an average weight of 18-20 g In preliminary experiments on white mice were determined by the minimum lethal dose of cultures of E. coli and S. cholerae suis. The minimum LD100for E. coli was 500 thousand microbial bodies (M.L.) in 1 ml, and for S. suis - 400 thousand M.L.

Conducted two series of tests of two samples of the integrated product, prepared as described in example otlichayushchayasya acid and levamisole. Each sample was tested in two versions: with prolongator PEG without PEG.

The first series. This series was tested the drug, including: 1% succinic acid; 2% of levamisole; 0.3% formalin; 7% of polyethylene glycol (PEG) and without PEG. The drugs were administered intraperitoneally in a volume of 0.25 ml.

The procedure of the experiment was the following. Mice of the first group of trial medication (without PEG) in a volume of 0.25 ml was introduced for 6 hours before infection with E.coli culture. The mice of the second group were injected similarly, the test drug with PEG and after 6 hours spent infection. Mice of the third and fourth groups were first infected with E.coli culture. After 6 hours the mice 3 groups was introduced trial medication without PEG, and mice 4 groups - the test drug with PEG. Control group mice were injected with saline for 3 hours before infection. The results of the study (Protective activity immunometabolic drug when modeling deadly infectious process culture of E. coli in a dose of 1 LD100) are presented in table 1.

Observation of the experimental mice was carried out within three days. During this time period, all mice in the control group fell. At autopsy, they were revealed changes characteristic of acute septic process.

When observation of the mice that were treated with the drug in combination with PEG showed h is about the survival rate was significantly higher. So, in group 4, the survival rate amounted to 85.7% versus 42.9% in relation to animals 3 groups.

The results of the first experimental experience testified to the fact that the metabolic composition has a certain protective effect when knowingly infecting mice with a lethal dose. Protective effect in long periods from infection significantly increased when the drug PEG.

The second series. This series was tested in the drug composition, comprising: a 1.5% succinic acid; 3% of levamisole; 0.3% formalin; 7% of polyethylene glycol (PEG) and without PEG. The drugs were administered intraperitoneally in a volume of 0.25 ml for 6 hours before infection. The results of the research (Protective metabolic activity of the drug in the simulation of mixed infection in white mice by culture of E.coli (2LD50and S. cholerae suis (2LD50) are presented in table 2.

Obtained in two series, the results indicate that the test drug has a definite anti-infective action. While anti-infective activity is expressed increases in distant time periods after infection, when included in the composition of the drug PEG.

The study of metabolic activity

In this series of experiments was used immunometabolic the drug, including: 1% succinic acid; 2% of levamisole; 0.3%formalin; 7% of polyethylene glycol and without PEG. The object of the experiments was suckling piglets-lipotropic 6-7 days of age, had a body weight of up to 1000

In accordance with the principle of analogs selected for the experiment pigs were divided into 2 groups. The piglets of the experimental group (n=8) was injected intramuscularly immunometabolic composition without PEG with multiplicity every 7 days. The amount of administered drug was 1.0 ml - 1,5-2,0-2,5 g In pigs of the second group (n=8) in the same manner and amount used the drug with PEG.

When the clinical observation found that already at day 2 after the introduction of the state of Guinea pigs of both experimental groups was significantly different from their peers in the control group. First, they became more cheerful; secondly, they have become more pronounced nutritional needs. These differences in piglets from the experimental groups relative to individuals from the control group increased with each passing day.

Study of the effect on hematological and immunobiochemical state of Guinea pigs spent on 7-14-21 and 28 days. The research results are reflected in table 3.

Based on the experience and results of the study for Hematology and immunobiochemical state of Guinea pigs made the following conclusion.

Parenteral administration of the test drug on the pigs-hypotrophic the x provides a pronounced stimulation of metabolic processes, what a favorable effect on the acceleration of their physiological development and the elimination of state of immunodeficiency.

Table 1
GroupMedicationProcessingThe death of mice after (hours)Survival
12-2424-3848-72abs.%
1 (n=7)Immunometabolic drug without PEG6 hours before infection13-342,8
2 (n=7)Immunometabolic drug +PEG6 hours before infection12-467,0
3 (n=7)Immunometabolic the drug without the AG After 6 hours after infection22-342,9
4 (n=7)Immunometabolic drug +PEGAfter 6 hours after infection1685,7
5 (n=5)Saline3 hours before infection32--

td align="center"> 1 (n=10)
Table 2
GroupMedicationThe death of mice after (hours)Survival
12-2424-4848-7272-9898-120abs.%
Immunometabolic drug without PEG212-550
2 (n=10)Immunometabolic drug +PEG12770
3 (n=5)Phys. solution21200

1. The method of obtaining comprehensive immunometabolic drug with anti-infective activity, including dissolved in demineralised water succinic acid, adding formalin solution, characterized in that as immunostimulant use levamisole in the following ratio, wt.%:

Succinic acid 1,0-1,5
Levamisole2,0-3,0
Formalin0,3-0,4
Distilled waterRest

2. The method of obtaining comprehensive immunometabolic drug with anti-infective activity, including dissolved in demineralised water succinic acid, adding formalin solution, characterized in that as immunostimulant use levamisole and additionally enter the glycol in the following ratio, wt.%:

Succinic acid1,0-1,5
Levamisole2,0-3,0
Formalin0,3-0,4
The polyethylene glycol7-8
Distilled waterRest



 

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(1)

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